Structure of chromatin from the pigeon brain. I. Composition, electrophoretic motility and circular dichroism spectra of nucleosome particles

Composition and structural properties of pigeon brain hemisphere neuron chromatin have been studied. In these cells nucleosomal DNA repeat length is about 165 nucleotide pairs. The content of H1 and H2A histones was found to decrease by 18 and 30% respectively in comparison with the chromatin possessing the normal quantity of those histones. At the same time the content of protein uH2A (A-24), being the conjugate of H2A histone and ubiquitine, is increased. Mononucleosomes isolated from neuron chromatin was found to have relatively low electrophoretic mobility in polyacrylamide gel taking into account the size of their DNA fragment. Circular dichroism spectra of nucleosome particles show that the neuron mononucleosomes are more unfolded than the rat thymus ones. Data obtained allow to suggest that the short DNA repeat and accumulation of protein uH2A in neurons are the factors influencing the compactization of neuron chromatin.     

Low complex I content explains the low hydrogen peroxide production rate of heart mitochondria from the long-lived pigeon, Columba livia

Across a range of vertebrate species, it is known that there is a negative association between maximum lifespan and mitochondrial hydrogen peroxide production. In this report, we investigate the underlying biochemical basis of the low hydrogen peroxide production rate of heart mitochondria from a long-lived species (pigeon) compared with a short-lived species with similar body mass (rat). The difference in hydrogen peroxide efflux rate was not explained by differences in either superoxide dismutase activity or hydrogen peroxide removal capacity. During succinate oxidation, the difference in hydrogen peroxide production rate between the species was localized to the ΔpH-sensitive superoxide producing site within complex I. Mitochondrial ΔpH was significantly lower in pigeon mitochondria compared with rat, but this difference in ΔpH was not great enough to explain the lower hydrogen peroxide production rate. As judged by mitochondrial flavin mononucleotide content and blue native polyacrylamide gel electrophoresis, pigeon mitochondria contained less complex I than rat mitochondria. Recalculation revealed that the rates of hydrogen peroxide production per molecule of complex I were the same in rat and pigeon. We conclude that mitochondria from the long-lived pigeon display low rates of hydrogen peroxide production because they have low levels of complex I.     

Absolute rates of adenosine formation during ischaemia in rat and pigeon hearts.

1. The activities of ecto- and cytosolic 5'-nucleotidase (EC 3.1.3.5), adenosine kinase (EC 2.7.1.20), adenosine deaminase (EC 3.5.4.4) and AMP deaminase (EC 3.5.4.6) were compared in ventricular myocardium from man, rats, rabbits, guinea pigs, pigeons and turtles. The most striking variation was in the activity of the ecto-5'-nucleotidase, which was 20 times less active in rabbit heart and 300 times less active in pigeon heart than in rat heart. The cytochemical distribution of ecto-5'-nucleotidase was also highly variable between species. 2. Adenosine formation was quantified in pigeon and rat ventricular myocardium in the presence of inhibitors of adenosine kinase and adenosine deaminase. 3. Both adenosine formation rates and the proportion of ATP catabolized to adenosine were greatest during the first 2 min of total ischaemia at 37 degrees C. Adenosine formation rates were 410 +/- 40 nmol/min per g wet wt. in pigeon hearts and 470 +/- 60 nmol/min per g wet wt. in rat hearts. Formation of adenosine accounted for 46% of ATP plus ADP broken down in pigeon hearts and 88% in rat hearts. 4. The data show that, in both pigeon and rat hearts, adenosine is the major catabolite of ATP in the early stages of normothermic myocardial ischaemia. The activity of ecto-5'-nucleotidase in pigeon ventricle (16 +/- 4 nmol/min per g wet wt.) was insufficient to account for adenosine formation, indicating the existence of an alternative catabolic pathway.     

Cisplatin: nephrotoxic action in vertebrates and its prevention

1. The kidney of frog and black sculpin appeared to be much less sensitive to the toxic action of CP in comparison with rat and pigeon. 2. The impairment of renal function after CP administration resulted in increased serum urea in rat, uric acid in pigeon and magnesium in black sculpin. 3. Kidney swelling is important feature of CP nephrotoxicity in rat and pigeon but not in frog and fish. 4. Pretreatment with choline chloride, PAH, furosemide and ethacrynic acid reduced the nephrotoxic action of CP in rat but did not prevent the accumulation of platinum in renal tissue which appeared to be a function of the dose injected to investigated animals.     

Arginine vasotocin induces free fatty acid release from avian adipose tissue in vitro

The effect of arginine vasotocin (AVT) on free fatty acid (FFA) release from pigeon adipose tissue slices incubated in vitro was studied. AVT at concentrations of 0.5 microgram/ml and 5.0 micrograms/ml was found to produce significant increases in the release of FFA from the adipose tissue. This augmentation of FFA release did not require the presence of hydrocortisone.     

The avian hippocampus, homing in pigeons and the memory representation of large-scale space

The extraordinary navigational ability of homing pigeons providesa unique spatial cognitive system to investigate how the brainis able to represent past experiences as memory. In this paper,we first summarize a large body of lesion data in an attemptto characterize the role of the avian hippocampal formation(HF) in homing. What emerges from this analysis is the criticalimportance of HF for the learning of map-like, spatial representationsof environmental stimuli used for navigation. We then exploresome interesting properties of the homing pigeon HF, using fordiscussion the notion that the homing pigeon HF likely displayssome anatomical or physiological specialization(s), comparedto the laboratory rat, that account for its participation inhoming and the representation of large-scale, environmentalspace. Discussed are the internal connectivity among HF subdivisions,the occurrence of neurogenesis, the presence of rhythmic thetaactivity and the electrophysiological profile of HF neurons.Comparing the characteristics of the homing pigeon HF with thehippocampus of the laboratory rat, two opposing perspectivescan be supported. On the one hand, one could emphasize the subtledifferences in the properties of the homing pigeon HF as possibledeparture points for exploring how the homing pigeon HF maybe adapted for homing and the representation of large-scalespace. Alternatively, one could emphasize the similarities withthe rat hippocampus and suggest that, if homing pigeons representspace in a way different from rats, then the neural specializationsthat would account for the difference must lie outside HF. Onlyfuture research will determine which of these two perspectivesoffers a better approximation of the truth.     

Non-enzymatic lipid peroxidation of microsomes and mitochondria isolated from liver and heart of pigeon and rat

Studies were carried out to determine the level of ascorbate-Fe2+ dependent lipid peroxidation of mitochondria and microsomes isolated from liver and heart of rat and pigeon. Measurements of chemiluminescence indicate that the lipid peroxidation process was more effective in mitochondria and microsomes from rat liver than in the same organelles obtained from pigeon. In both mitochondria and microsomes from liver of both species a significant decrease of arachidonic acid was observed during peroxidation. The rate C18:2 n6/C20:4 n6 was 4.5 times higher in pigeon than in rat liver. This observation can explain the differences noted when light emission and unsaturation index of both species were analysed. A significant decrease of C18:2 n6 and C20:4 n6 in pigeon liver mitochondria was observed when compared with native organelles whereas in pigeon liver microsomes only C20:4 n6 diminished. In rat liver mitochondria only arachidonic acid C20:4 n6 showed a significant decrease whereas in rat liver microsomes C20:4 n6 and C22:6 n3 decreased significantly. However changes were not observed in the fatty acid profile of mitochondria and microsomes isolated from pigeon heart. In the heart under our peroxidation conditions the fatty acid profile does not appear to be responsible for the different susceptibility to the lipid peroxidation process. The lack of a relationship between fatty acid unsaturation and sensitivity to peroxidation observed in heart suggest that other factor/s may be involved in the protection to lipid peroxidation in microsomes and mitochondria isolated from heart.     

Potential use of exogenous RNAs for the analysis of the complex effects of biologically active substances

It has been established in rat experiments that RNA obtained from the liver and renal cortex of animals given hydrocortisone produces in recipients the rise of physical endurance evoked by the hormone. RNA obtained from other organs of these animals and from any test organs of control donors did not influence physical endurance. The key role of the liver and kidneys in the realization of the hydrocortisone effect is likely to be connected with activation of the synthesis of gluconeogenesis enzymes. RNA obtained from the organs of donors premedicated with hydrocortisone reproduced stimulation of gluconeogenesis with hydrocortisone. The use of exogenous RNA holds promise for analysis of complex effects of biologically active substances, since it permits studying separately the components of the effects determined by activation of protein synthesis in definite organs.     

Enhancement of α-lactalbumin-like activity in mammary explants from pregnant rats in the absence of exogenous prolactin

Mammary explants from pregnant rats showed a progressive increase in α-lactalbumin activity during culture with insulin, hydrocortisone and prolactin. Unexpectedly, culture with only insulin and hydrocortisone produced a similar rate of increase of α-lactalbumin-like activity, but this increase commenced about 24 hr later. The delay suggests that the enhanced activity effected by insulin and hydrocortisone is not a reflection of carry-over of endogenous mammotrophic hormones. Insulin plus hydrocortisone did not stimulate casein or fatty acid synthesis by pregnancy tissue, and did not enhance α-lactalbumin-like activity in virgin rat mammary explants. Enhancement of this activity by insulin plus hydrocortisone in pregnant tissue was constant over a wide range of glucocorticoid concentrations, but was inhibited by progesterone. Available evidence indicates that the active factor in extracts from insulin-hydrocortisone-explants is a heat-stable protein which is either α-lactalbumin itself, or another molecule with similar specifier properties.     

Immunohistochemical and quantitative analysis of 5-hydroxytryptamine in the retina of some vertebrates

5-Hydroxytryptamine immunoreactive neurons were found in retinae from chicken, pigeon, frog and goldfish. They were localized among the amacrine cells with a distribution of cell bodies and nerve fibres that varied with the species. In chicken and pigeon, bipolar-like cell bodies were also found in the middle of the inner nuclear layer, sending processes inwards to the inner plexiform layer and outwards to the horizontal cells. The signalling direction of these cells is doubtful. No 5-hydroxytryptamine immunoreactivity was found in retinae from cow, pig, cat, rabbit, guinea-pig, rat or mouse.Quantitative analyses were performed with HPLC on extracts from chicken, pigeon, frog and goldfish retinae. High concentrations were found in goldfish and frog whereas less, about 100 ng/g, was observed in chicken and pigeon.The results suggest that 5-hydroxytryptamine is the transmitter of a set of amacrine cells in cold-blooded vertebrates and perhaps also in birds. The transmitter of the indoleamine accumulating neurons of mammals remains to be further elucidated.     

Effect of dimethoate on hepatic cytochrome P-450 and glutathione S-transferase activity in pigeon and rat

Effect of acute exposure (24 hr) to different oral doses of dimethoate on hepatic microsomal cytochrome P-450 (Cyt. P-450) content and cytosolic glutathione S-transferase (GST) activity were determined in pigeon and rat to ascertain difference in the metabolic response as a measure of species selective toxicity. Dimethoate at five different doses caused a statistically significant decrease in Cyt. P-450 content both in pigeon and rat. However, reduction in GST activity was significant at three doses in pigeon and at high dose in rat. Thus, a different quantum of hepatic Cyt. P-450 decrease and a differed response of GST activity against dimethoate exposure in pigeon and rat may be one of the possible causes for relatively higher toxicity of dimethoate in birds.     

Comparative aspects of aminotransferases in the rat, pigeon and rainbow trout.

1. The activities of aminotransferases catalysing the transfer of amino groups from aspartate, alanine and leucine to 2-oxoglutarate in different tissues of the rat, pigeon and trout have been determined. 2. Alanine-2-oxoglutarate aminotransferase was high in the liver of the rat and trout and low in that of the pigeon. 3. Aspartate-2-oxoglutarate aminotransferase was usually the dominant aminotransferase in all tissues and was highest in oxidative tissues where the TCA cycle is active. Its activity in the various livers is not correlated with the function of aspartate in nitrogen excretion. 4. The activity of aspartate-2-oxoglutarate aminotransferase in oxidative tissues argues that aspartate in conjunction with this enzyme serves as a buffer of oxaloacetate to keep the TCA cycle running and/or to mediate the transfer of reducing equivalents across mitochondrial membranes.     

Age-related differential induction of tryptophan pyrrolase by hydrocortisone in the liver of male rats

The activity and the regulatory pattern of tryptophan pyrrolase of the liver of male rats during various phases of the life span were studied with a view to investigate the differential effectiveness of hydrocortisone in relation to growth, development, and senescence of an organism. The level of this enzyme shows no significant change till adulthood but decreases significantly thereafter with increasing age. Adrenalectomy and hydrocortisone treatments decrease and increase, respectively the activity of this enzyme significantly in rats of all the ages. However, the effects of these treatments are highest in the mature rat. Induction of the enzyme by hydrocortisone is actinomycin D-sensitive.     

Effects of adrenalectomy and hydrocortisone on DNA synthesis in the rat anterior pituitary gland

The effect of adrenalectomy and hydrocortisone on /³H/-thymidine uptake by rat anterior pituitary cells in the short-time organ culture was investigated. Adrenalectomy significantly increases the incorporation tritiated thymidine into anterior pituitary cells nuclei, whereas hydrocortisone has an opposit effect. The existence on negative feedback between the intracellular hormone content and DNA synthesis in ACTH-secreting anterior pituitary cells is suggested.     

Regulation of glucosamine synthetase activity by cholesterol and hydrocortisone

The effects of cholesterol and hydrocortisone (cortisol) on the activity of purified glucosamine synthetase from rat liver was studied in vitro. It was found that the enzyme activity is increased by cholesterol and inhibited by hydrocortisone. These steroids block the allosteric effect of vitamin K1 on the enzyme. There is evidence testifying to the allosteric type of cholesterol and hydrocortisone effects on glucosamine synthetase.     

The relationship between mitochondrial heterogeneity and gluconeogenesis in liver mitochondria of the rat, pigeon and guinea pig.

Isolated mitochondria of pigeon and guinea pig liver were subjected to zonal centrifugation. With pigeon liver mitochondria there was uniform distribution of pyruvate carboxylase, phosphoenolpyruvate carboxykinase, malate dehydrogenase, aspartate aminotransferase and glutamate dehydrogenase activities. Guinea pig liver mitochondria demonstrated two pyruvate carboxylase and phosphoenolpyruvate carboxykinase maxima but only one maximum with aspartate aminotransferase, malate dehydrogenase and glutamate dehydrogenase. Mitochondrial enzyme levels in rat, pigeon and guinea pig indicate different roles of certain gluconeogenic enzymes in the transport of carbon and hydrogen in and out of mitochondria.     

The effects of the ionophore A-23187 on the rat vas deferens

The calcium ionophore A-23187 induced spontaneous, rhythmic contractions in the rat isolated vas deferens in a concentration-dependent manner. Contractions were blocked by phentolamine and were abolished following pretreatment with reserpine. In tissues preloaded with [3H]noradrenaline, A-23187 (10 microM) caused a time-dependent increase in the release of tritium. The findings suggest that A-23187-induced contractions in the rat vas deferens are secondary to the release of endogenous noradrenaline from the adrenergic nerves, as are contractions induced in this preparation by X-537A (another calcium ionophore) described earlier by other investigators.     

Nuclear localization of gold labeled-hydrocortisone-bovine serum albumin conjugate injected intravenously into the hormone-target cells of rat.

We have suggested in a previous study using 2-nm colloidal gold labeled-testosterone-bovine serum albumin (testosterone-BSA-gold) that 2-nm gold labeled-steroid hormone-BSA conjugates would be a useful tool for analyzing the mechanism of steroid hormone action (39). In this study, we examined whether hydrocortisone-BSA conjugate (hydrocortisone-BSA) showed a similar distribution to radiolabeled hydrocortisone in vivo, by injecting 2-nm colloidal gold labeled-hydrocortisone-BSA (hydrocortisone-BSA-gold) into the rat tail vein. The hydrocortisone-BSA-gold with silver enhancement became visible as silver deposits under electron microscopy in the nuclei of hepatocytes and hepatic stellate cells but not in Kupffer cells in the liver, and in the thymocytes and thymic reticuloepithelial cells in the thymus of a rat killed 2 h postinjection. The percentage of nuclei showing deposits in the non-target cells, the epithelial cells of the seminal vesicle, was similar to the value in the seminal vesicle of a control rat injected with BSA labeled with 2-nm colloidal gold as reported previously. In the hepatocytes and thymocytes of a control rat not injected, the percentages of nuclei showing deposits were similar to those in the rat injected with testosterone-BSA-gold or BSA-gold as reported previously, but lower than those in the rat injected with hydrocortisone-BSA-gold. These results suggest that hydrocortisone-BSA-gold is useful for the morphological study of hydrocortisone target cells, and imply that BSA conjugated with hydrocortisone can enter the target cell nuclei of the rat. The present study further indicates that the fate of gold labeled-steroid hormone-BSA conjugates may be decided at the cell membrane level.     

Alterations of pancreatic juice amylase by glucocorticoid levels in the rat

By use of isoelectrofocusing, three isoenzymes with pIs of 8.40, 8.55, and 8.65 were characterized in the amylase fraction of rat pancreatic juice. Enzyme secretion in rat exocrine pancreas is affected by glucocorticoid levels; adrenalectomy led to a significant decrease in protein secretion which was more pronounced in the amylase fraction, in which the isoenzymes with pI 8.55 and 8.65 disappeared. Substitution therapy with hydrocortisone (25 mg/kg/day, for 6 days) restored exocrine pancreatic secretion to almost normal levels. Administration of hydrocortisone to control rats led to structural alterations in enzymes secreted, splitting the amylase isoenzymes with pI 8.40; this was confirmed by crossed immunoelectrophoresis. It is concluded that glucocorticoid levels play an important role in the maintenance of function of exocrine pancreas and it is suggested that, although hydrocortisone fulfills the objective of restoring enzyme secretion diminished by adrenalectomy, it is possible that intensive treatment could have undesirable effects on the structure of enzymes and could involve pancreatic disfunctionality.